Hepatitis C virus is the major cause of transfusion and community-acquired non-A, non-B hepatitis worldwide. Approximately 2% of the world's population are infected with the virus. In the Unites States, hepatitis C represents approximately 20% of cases of acute hepatitis. Unfortunately, self-limited hepatitis is not the most common course of acute HCV infection. In the majority of patients, symptoms of acute hepatitis resolve, but alanine aminotransferase (a liver enzyme diagnostic for liver damage) levels often remain elevated and HCV RNA persists. Indeed, a propensity to chronicity is the most distinguishing characteristic of hepatitis C, occurring in at least 85% of patients with acute HCV infection. The factors that lead to chronicity in hepatitis C are not well defined. Chronic HCV infection is associated with increased incidence of liver cirrhosis and liver cancer. No vaccines are available for this virus, and current treatment is restricted to the use of alpha interferon, which is effective in only 15-20% of patients. Recent clinical studies have shown that combination therapy of alpha interferon and ribavirin leads to sustained efficacy in 40% of patients (Poynard, T. et al. Lancet (1998), 352, 1426-1432.). However, a majority of patients still either fail to respond or relapse after completion of therapy. Thus, there is a clear need to develop more effective therapeutics for treatment of HCV-associated hepatitis.
HCV is a positive-stranded RNA virus. Based on comparison of deduced amino acid sequence and the extensive similarity in the 5′ untranslated region, HCV has been classified as a separate genus in the Flaviviridae family, which also includes flaviviruses such as yellow fever virus and animal pestiviruses like bovine viral diarrhea virus and swine fever virus. All members of the Flaviviridae family have enveloped virions that contain a positive stranded RNA genome encoding all known virus-specific proteins via translation of a single, uninterrupted, open reading frame.
Considerable heterogeneity is found within the nucleotide and encoded amino acid sequence throughout the HCV genome. At least six major genotypes have been characterized, and more than 50 subtypes have been described. The major genotypes of HCV differ in their distribution worldwide, and the clinical significance of the genetic heterogeneity of HCV remains elusive despite numerous studies of the possible effect of genotypes on pathogenesis and therapy.
The RNA genome is about 9.6 Kb in length, and encodes a single polypeptide of about 3000 amino acids. The 5′ untranslated region contains an internal ribosome entry site (IRES), which directs cellular ribosomes to the correct AUG for initiation of translation. As was determined by transient expression of cloned HCV cDNAs, the precursor protein is cotranslationally and posttranslationally processed into at least 10 viral structural and nonstructural (NS) proteins by the action of a host signal peptidase and by two distinct viral proteinase activities. The translated product contains the following proteins: core-E1-E2-p7-NS2-NS3-NS4A-NS4B-NS5A-NS5B.
The N-terminal portion of NS3 functions as a proteolytic enzyme that is responsible for the cleavage of sites liberating the nonstructural proteins NS4A, NS4B, NS5A, and NS5B. NS3 has further been shown to be a serine protease. Although the functions of the NS proteins are not completely defined, it is known that NS4A is a protease cofactor and NS5B is an RNA polymerase involved in viral replication. Thus, agents that inhibit NS3 proteolytic processing of the viral polyprotein are expected to have antiviral activity.
Extensive efforts toward the development of HCV NS3 protease inhibitors have resulted in the following disclosures: WO 98/17679 (Tung et al.) describes a large class of generic peptide and peptidomimetic inhibitors with the following formula: U-E8-E7-E6-E5-E4-NH—CH(CH2G1)-W1, wherein W1 is a variety of electrophilic groups. E4 represents either an amino acid or one of a series of peptidomimetic groups. No example of compounds wherein W1 is boronic acid or ester is disclosed or enabled in WO 98/17679. Additionally, compounds with extended aralkyl or heteroaralkyl P1 substituents as disclosed in the present application are not disclosed, enabled or exemplified in WO 98/17679.
WO 98/22496 (Attwood et al.) discloses solely hexapeptide inhibitors of the following general formula: R9—NH—CH(R8)—CO—NH—CH(R7)—CO—N(R6)—CH(R5)—CO—NH—CH(R4)—CO—N(R3)—CH(R2)—CO—NH—CH(R1)-E wherein E is either an aldehyde or a boronic acid. Compounds with extended aralkyl or heteroaralkyl P1 substituents as disclosed in the present application are not specifically disclosed, enabled or exemplified in WO 98/22496.
WO 99/07734 (Llinas-Brunet et al.) discloses tetra- to hexa-peptide analogs containing a P1 electrophilic carbonyl group, a phosphonate ester, or an aza-aminoacid analog. WO 99/07733 (Llinas-Brunet et al.) describes related peptides terminating in a carboxylate. Similar compounds are reported by Steinkuhler et al. Biochemistry (1998), 37, 8899-8905 and Ingallinella et al. Biochemistry (1998), 37, 8906-8914. None of these publiscations teaches the making and use of compounds with aralkyl or heteroaralkyl P1 substituents.
WO 99/50230 (Tung et al.) discloses peptidomimetics containing a 5 or 6-membered carbocyclic ring at the P2 position. Tung et al. does not teach the aralkyl or heteroaralkyl P1 substituents of the present invention.
WO 00/09543 (Llinas-Brunet et al.) discloses tripeptides containing a substituted proline residue at P2 and an aminocyclopropanecarboxylate derivative at P1. A related disclosure, WO 00/09558 (Llinas-Brunet et al.), discloses tetra- to hexapeptides with the same P1 and P2 structure as WO 00/09543.
Other peptide inhibitors of HCV protease have been disclosed. WO 98/46630 (Hart et al.) has described hepta-peptide analogs containing an ester linkage at the scissile bond. WO 97/43310 (Zhang et al.) discloses high molecular weight peptide inhibitors. The present invention is distinct from the compounds of WO 98/46630 or WO 97/43310.
Additionally, literature regarding HCV NS3 protease inhibitors suggest that the S1 pocket of the NS3 protease enzyme can only accommodate small aliphatic P1 residues. (Pizzi et al. Proc. Natl. Acad. Sci. USA (1994), 91, 888-892; Urbani et al. J. Biol. Chem. (1997) 272, 9204-9209; Perni, Robert B. Drug News Perspective (2000), 13, 69-77). Thus, the general literature regarding HCV NS3 protease inhibitors does not suggest or provide the motivation to one skilled in the art to make extended aralkyl P1 inhibitors of the present invention.
Based on the large number of persons currently infected with HCV and the limited treatments available, it is desirable to discover new inhibitors of HCV NS3 protease. The instant invention discloses a class of novel peptides with extended P1 residues that exhibit inhibitory activity against HCV NS3 protease. Further, the present invention discloses unexpected benefit of HCV NS3 protease inhibitory selectivity over inhibition of elastase and/or chymotrypsin.